We investigated seawater microbial abundance, activity and diversity in a site strongly influenced by submarine groundwater discharge (SGD). Three sampling campaigns (August 2020, February 2021 and July 2021) were conducted at a field site, highly influenced by SGD (Achziv, northern Israel), Each field campaign lasted 2-5 days and covered at least 2 tidal cycles. Pore-water samples were collected on the shoreline using piezometers (AMS piezometers that reach depths of <2 meters) and a portable peristaltic pump. The density (g cm-3), electric conductivity (mS/cm), temperature (°C) and pH, of surface seawater, porewater and groundwater were measured on-site at the time of the sampling. Samples for microbial analysis were collected from the piezometers and divided to aliquots: 1. For community analysis, samples were immediately filtered through polycarbonate 0.2 μm pore size filters, which were kept on ice and transported to the laboratory on the same day. Filter samples were stored frozen (-20°C) until DNA extraction (filtered pore-water were kept for dissolved nutrient measurements. After thawing, each filter was cut into small pieces using a sterile scalpel blade, which was placed immediately into PowerSoil DNA bead tubes and extracted with the dNeasy PowerSoil Kit (Qiagen, USA) following the standard protocol. 2. For Pico-/nano-phytoplankton and heterotrophic prokaryotic abundance, non-filtered samples were chilled on ice and transported to the laboratory on the same day. Samples (1.8 mL) were fixed with glutaraldehyde (final concentration 0.02 % v:v, Sigma-Aldrich 253 G7651), frozen in liquid nitrogen, and later stored at −80°C until analysis. The abundance of autotrophic pico- and nano-eukaryotes, Synechococcus and Prochlorococcus, and other heterotrophic prokaryotes (bacteria and archaea) was determined using an Attune® Acoustic Focusing Flow Cytometer (Applied Biosystems) equipped with a syringe based fluidic system and 488 and 405 nm lasers. To measure heterotrophic prokaryote abundance, a sample aliquot was stained with SYBR Green (Applied Biosystems). 3. Prokaryotic (bacteria and archaea) heterotrophic production was estimated using the 3H-leucine incorporation method. Photosynthetic carbon fixation rates were estimated using the 14C incorporation method.
